P462: RECIPIENT SEX AND DONOR RISK GROUP INFLUENCE LEUKEMIA KINETICS IN NSG MICE
Authors: Marlon Arnone, Anna Stanger, Pauline Hanns, Jessica Kuebler, Elsa Goersch, Saskia Rudat, Claudia Lengerke
Journal: HemaSphere (2023)

Background: Healthy hematopoietic stem and progenitor cells and acute myeloid leukemia (AML) cells heavily rely on interactions with their environment and fine-tuned extrinsic signaling. As a result, ex vivo expansion and functional in vitro studies with primary AML cells remain challenging. Patient-derived xenograft (PDX) models can be used to study human cells in vivo, however a significant number of primary AML samples were reported as not engraftable in this model. Aims: We provide a detailed characterization of PDX models in NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice to elucidate variables influencing the kinetics of leukemia development and expansion in vivo and inform further functional studies. Methods: 60 AML cases of all ELN risk categories were transplanted intra-femorally into sub-lethally irradiated NSG mice. To assess a potential impact of recipient sex, AML cells from the same patient were transplanted side-by-side in male and female recipient mice. Animals were routinely screened using flow cytometry for leukemic cells in bone marrow (BM) at weeks 16, 26 or 39 post-transplantation and monitored for overall survival (OS). To evaluate homing ability, 1 million primary AML cells were labelled with CFSE and injected intra-venously without prior irradiation. BM and peripheral blood were collected 16 hours post-injection and analyzed by flow cytometry for CFSE-positive AML cells. Results: Interestingly, AML cells transplanted into female recipients showed much higher engraftment than cells from the same AML case transplanted into male NSG recipients (70.8% vs. 27.91% leukemic cells among total BM cells) when analyzed at the same time-point post-transplantation. Secondary transplants reproduced this effect showing 58.7% vs. 1.77% leukemic cells among total BM cells in female vs. male recipients transplanted with leukemic cells from the same source. Based on these data, sex-matched recipients were used for the subsequent engraftment of our cohort. Adverse risk AML cells associated with a shorter OS (31.5 weeks) than favorable (53 weeks) or intermediate (39 weeks) AML samples. High FLT3-internal tandem duplication (ITD) mutational burden (high ITD/wild-type ratio) was a negative prognostic factor with regards to murine OS, when compared to FLT3 wild-type (WT) (28 vs 48 weeks). Animals transplanted with FLT3-ITD/NPM1 double mutant cells showed shorter OS (28 weeks) compared to those receiving WT cells (52 weeks) or single NPM1 mutant cells (41.5 weeks). FLT3-ITD single mutant cells further induced shorter OS (38 weeks) compared to WT cells. Mice transplanted with samples from patients that did or did not achieve complete remission after first induction displayed different OS (42.5 vs 32 weeks, p= 0.2). AML cell morphology (FAB classification) and phenotype also influenced murine OS. Given the close association between risk factors and outcome in humans and mice, we asked whether human AML cell homing capacity in NSG mice could be established as a short-term assay predicting disease aggressiveness in patients. Indeed, higher homing rates were observed for severe and intermediate risk groups vs favorable AML samples (0.0085% and 0.0075% vs 0.0059%, respectively) but no predictive value was found with respect to patient or murine OS. Summary/Conclusion: We conclude that several features of human AML are faithfully recapitulated in PDX models and that these represent a robust model for in vivo functional studies on primary human AML cells. Homing, as measured by our current assay, may not be a reliable predictor for long-term outcome perhaps because it also captures cells that do not lead to leukemia induction. Recipient sex is an important unreported parameter that needs to be considered for all experiments in PDX models. Keywords: Mouse model, Acute myeloid leukemia, Survival prediction